The entry of an enveloped virus into a target cell is initiated by the envelope protein of the virus binding to a receptor on the target cell membrane. The interaction of the receptor and the viral envelope protein triggers fusion between the envelope protein and the cell membrane, permitting the entry of the viral genome into the cytoplasm of the cell. The fusion mediated by the viral envelope protein also permits cell-to-cell transmission of enveloped viruses. The expression of the viral envelope protein on the membrane of an infected cell can trigger fusion with an uninfected cell bearing the appropriate viral envelope protein receptor, further increasing the virulence of the virus within the host. Perhaps the most well-known example of the contribution of viral-mediated cell fusion to a disease state is the viral-induced fusion that results in the formation of multinucleated giant cells (synctia) after human immunodeficiency virus (HIV) infection, which eventually leads to death of the fused cells. Thus, the receptor-viral envelope protein determines the tropism, virulence, and ultimately the pathogenicity of the virus in a particular host.
Structural, biochemical, and functional studies have revealed a number of common features for viral membrane fusion proteins. At least two groups of fusion proteins have been identified to date. Dutch, R. E., et al., Biosci. Rep. 20: 597-612 (2000). In one group, the fusion proteins that include the paramyxovirus F protein, the HIV gp 160 protein, the HTLV SU protein, the Ebola GP protein, and the influenza hemagglutin (HA) protein. These viral envelope proteins share common features that include multiple glycosylation sites, a trimeric structure, and proteolytic cleavage for fusogenic activation. For each of the fusion proteins, the proteolytic cleavage resulted in an extremely hydrophobic subunit near the new N-terminus identified as the fusion peptide. Three heptad repeats were identified near the fusion peptide and the transmembrane domain that formed a trimeric coiled coil. Therefore, within widely disparate groups of enveloped viruses, a common mechanism of viral membrane fusion appears to function using the trimeric coiled coil motif. In a second group that includes togavirus, rhabodovirus, and flavivirus, fusion appears to occur by another molecular mechanism.
Membrane fusion events may be mediated by interaction of the viral envelope protein with one or more receptors on a target cell. For example, a single protein serves both as the viral cell recognition and fusion proteins (e.g., influenza) whereas in other viruses, these activities are separated (e.g., HIV). Alternately stated, the viral envelope protein can sometimes require both a receptor and a co-receptor for the initiation of fusion events. See, e.g., Overbaugh, et al., Microbiol. & Mol. Biol. Rev. 65:371-389 (2001). To date, only HIV has been shown to use co-receptors to mediate the fusogenic event. Additionally, membrane fusion may occur under pH-independent or pH-dependent conditions.
In one example of viral fusion, the entry of HIV into target cells is mediated by a fusion reaction in which the gp120/gp41 glycoprotein of the virus binds to CD4 and a CC chemokine receptor, CCR5 or CXCR4, on the target cell membrane. The HIV enveloped surface glycoproteins are synthesized as a single 160 kD precursor protein which is cleaved by a cellular protease during viral budding into two glycoproteins, gp41 and gp120. gp41 is a transmembrane protein and gp120 is an extracellular protein which remains non-covalently associated with gp41, possibly in a trimeric or multimneic form. Hammarskjold, M. et al., Biochem. Biophys. Acta 989:269-280 (1989). HIV is targeted to CD4+ cells because the CD4 cell surface protein acts as the cellular receptor for the HIV-I virus. See, e.g., Dalgleish, A. et at, Nature 312:763-767 (1984); Klatzmann et al., Nature 312:767-768 (1984). Viral entry into cells is dependent upon gp120 binding the cellular CD4+ receptor molecules. See (McDougal, J. S. et al., Science 231:382-385 (1986); Maddon, P. J. et al., Cell 47:333-348 (1986).
The tropism and virulence of HIV appears to be determined by the co-receptor used to bind the viral envelope protein. HIV strains using the CCR5 as a co-receptor mediate transmission and predominate early in the course of disease, while those using CXCR4 are mainly associated with the symptomatic phase. HIV pathogenesis is very sensitive to cell surface CCR5 levels. Individuals who are homozygous for the CCR5 null allele D32 are resistant to infection. Heterozygotes for this allele, although not protected from infection, experience a delay of about two years in the onset of disease symptoms. This is apparently a result of the rather modest 50% reduction in cell surface CCR5 compared to wild-type CCR5 cells.
Membrane fusion also occurs as part of the intracellular vesical machinery. These membrane processes permit the transport of material between cellular compartments and out of the cell. This fusion process is mediated by a set of conserved proteins collectively termed SNARES. Solinar et al., Nature 362: 318-24 (1993). The proteins mediating the fusogenic activity also have a coiled coil motif similar to that of viral envelope proteins. Sutton, et al., Nature 395: 347-53.